Improved understanding of cellular and molecular events which occur at the interface between tissues and implants is beginning to allow new approaches to biomaterial design. The challenge is to produce biomaterials that are engineered to elicit specific, clinically-desirable responses from living cells and tissues in a patient's body. For example, it is clinically desirable for osteoblasts to rapidly deposit mineralized matrix on the surface of (or in close apposition to) newly implanted prostheses. The swift deposition of bone stabilizes the prosthesis and minimizes motion-induced damage to surgically traumatized tissue at the implantation site.
Anchorage-dependent cells (such as osteoblasts) must first adhere to a surface in order to perform subsequent cellular functions (e.g., proliferation, deposition of bone tissue, etc.). Since cell adhesion is the key to subsequent events, methods for promoting cell adhesion are of considerable interest. The effects on cell adhesion of peptides immobilized on the surfaces of substrates have been reported. Substrates have included polymers [Massia and Hubbell Journal of Biomedical Materials 25, 223-242 (1991)] and dental/orthopedic implant materials such as Cobalt-Chromium-Molybdenum alloy [Mikos et al. Biomaterials for Cell and Drug Delivery 331, 269-274 (1994)]. Adhesion-related peptides that have been attached to substrates have been mainly integrin-binding peptides, such as those which contain the Arginine-Glycine-Aspartic Acid (RGD) sequence.
The present invention relates to novel, heparin-binding, osteoblast-adhesive amino acid sequences of the formula: EQU aa.sup.1 -aa.sup.2 -aa.sup.3 -aa.sup.4
wherein: PA1 wherein: PA1 Aa.sup.1 represents the residue of an amino acid selected from the group consisting of H-Lys, H-Arg, H-Orn, and 6-aminocaproic acid (Acp); PA1 aa.sup.2, and aa.sup.4 independently represent the residue of an amino acid selected from the group consisting of lysine (Lys), arginine (Arg), and ornithine (Orn); PA1 aa.sup.3 represents the residue of an amino acid selected from the group consisting of alanine (Ala), glycine (Gly), valine (Val), leucine (Leu), isoleucine (Ile), serine (Ser), threonine (Thr) cysteine (Cys), methionine (Met), asparagine (Asn), norleucine (Nle), norvaline (Nva), and 2-aminobutyric acid (Abu); and n is zero or an integer from 1 to 6. The invention also includes pharmaceutically acceptable salts of the foregoing compounds. Under certain circumstances, discussed below, tetrapeptides of the following formulae are preferred: H-Lys-Arg-Met-Arg-OH (n=0 SEQ ID NO:12); H-Lys-Arg-Ala-Arg-OH (n=0 SEQ ID NO:19); H-Arg-Arg-Ser-Arg-OH (n=0 SEQ ID NO:26); H-Orn-Arg-Ser-Arg-OH (n=0 SEQ ID NO:33); H-Lys-Lys-Ser-Lys-OH (n=0 SEQ ID NO:40); H-Acp-Arg-Ser-Arg-OH (n=0 SEQ ID NO:47). The peptide sequence which is most preferred is H-Lys-Arg-Ser-Arg-OH (i.e. n=0 SEQ ID NO:1). The single-letter code for Lys-Arg-Ser-Arg is KRSR.
aa.sup.1 represents the residue of an amino acid selected from the group consisting of H-Lys, H-Arg, H-Orn, and 6-aminocaproic acid (Acp); PA2 aa.sup.2, and aa.sup.4 independently represent the residue of an amino acid selected from the group consisting of lysine (Lys), arginine (Arg), and ornithine (Orn); and PA2 aa.sup.3 represents the residue of an amino acid selected from the group consisting of alanine (Ala), glycine (Gly), valine (Val), leucine (Leu), isoleucine (Ile), serine (Ser), threonine (Thr), cysteine (Cys), methionine (Met), asparagine (Asn), norleucine (Nle), norvaline (Nva), and 2-aminobutyric acid (Abu). This peptide sequence has not been previously shown to exhibit any biological activity.